Differentiation of human pluripotent stem cells to cells similar to cord-blood endothelial colony–forming cells

Abstract

The ability to differentiate human pluripotent stem cells into endothelial cells with properties of cord-blood endothelial colony–forming cells (CB-ECFCs) may enable the derivation of clinically relevant numbers of highly proliferative blood vessel–forming cells to restore endothelial function in patients with vascular disease. We describe a protocol to convert human induced pluripotent stem cells (hiPSCs) or embryonic stem cells (hESCs) into cells similar to CB-ECFCs at an efficiency of >108 ECFCs produced from each starting pluripotent stem cell. The CB-ECFC-like cells display a stable endothelial phenotype with high clonal proliferative potential and the capacity to form human vessels in mice and to repair the ischemic mouse retina and limb, and they lack teratoma formation potential. We identify Neuropilin-1 (NRP-1)-mediated activation of KDR signaling through VEGF165 as a critical mechanism for the emergence and maintenance of CB-ECFC-like cells.

CD44 expression in endothelial colony-forming cells regulates neurovascular trophic effect

“…ECFCs can be isolated from human cord or peripheral blood and have robust clonal proliferative potential. They have been reported to home to the site of tissue ischemia after intravenous injection, where they improve circulation in a model of myocardial infarction (4), stroke (5), ischemic retinopathy (6,7), and ischemic limb injury (8,9). Although a paracrine trophic rescue effect of ECFCs has been postulated (10,11), factors that may mediate this effect remain poorly characterized.…”

Generating induced pluripotent stem cell derived endothelial cells and induced endothelial cells for cardiovascular disease modelling and therapeutic angiogenesis

“…Alternatively, monolayer culture of hiPSCs on matrigel, in defined media, with timed addition of growth factors has been shown to produce endothelial cells at an efficiency of up to 30% [64,65]. Notably, Prasain et al differentiated iPSCs to cordblood endothelial colony forming cells (ECFCs), a population of circulating endothelial progenitor cells with much higher proliferative capacity [66]. They used a two-step protocol involving partial embryoid body formation and subsequent 2D cell culture and isolated cells positive for CD31 and NRP-1 (neuropilin-1), a co-receptor for VEGF expressed early on in endothelial lineage differentiation [66].…”

Evaluation of Nonperfused Retinal Vessels in Ischemic Retinopathy

“…Furthermore, cell therapy approaches, aiming to regenerate ischemic retinal vasculature 41,42 will depend on a persistent extracellular template. Thus, our finding of persistent basement membrane after occlusion, demonstrates that therapies aiming for reperfusion in ischemic eye disease are possible in principle.…”